[3][4][5] At the time, the scientific community was largely unaware of the importance of protein phosphorylation in the regulation of cellular processes, and many in the field dismissed phosphoproteins as biologically unimportant.

Since covalent modification by phosphorylation is a widespread, important method of biochemical regulation in a wide variety of cellular processes, the discovery of this reaction has had enormous impact on scientific understanding of regulatory mechanisms.

However, for unknown reasons at the time, the only way to isolate glycogen phosphorylase a from muscle tissue was by paper filtration – other methods, such as centrifugation, would not work.

It was a critical insight on the part of Fischer et al. that it was the presence of calcium ions in the filter paper that was generating the active "a" isoform.

[9][10][11][12][13] While this may seem surprising given that it was isolated over 50 years ago, there are significant difficulties in studying the finer details of PhK's structure and mechanism due to its large size and high degree of complexity.

[11][12] Structural and biochemical data suggest one possible mechanism of action for the phosphorylation of glycogen phosphorylase by PhK involves the direct transfer of phosphate from adenosine triphosphate (ATP) to the substrate serine.

[9] Phosphorylase kinase is a 1.3 MDa hexadecameric holoenzyme, though its size can vary somewhat due to substitution of different subunit isoforms via mRNA splicing.

Additionally, the release of calcium ions from the sarcoplasmic reticulum during muscle contraction inactivates the inhibitory δ subunit and activates PhK fully.

When the cell needs to stop glycogen breakdown, PhK is dephosphorylated by protein phosphatases 1 and 2, returning the α and β subunits to their initial inhibitory configuration.